The invention relates to the field of methods and probe designs for obtaining quantitative optical properties and chromophore concentrations of tissue components in-vivo at superficial depths and “short” source-detector separations.
The absorption coefficient μa, the scattering coefficient μs′, and chromophore concentrations of skin are fundamental properties of tissue that can provide essential information for many aesthetic, therapeutic, and diagnostic applications such as monitoring of skin blood oxygenation, melanin concentration, detection of cancer with fluorescence, laser surgery, and photodynamic therapy. Many researchers have quantified the optical properties of skin tissue and most of them used ex-vivo skin samples and integrating sphere techniques.
Although the integrating sphere based techniques can be used to investigate ex vivo optical properties of epidermis, dermis, and subcutaneous tissue, the need to take biopsies from subjects limits their applicability in the clinic. Furthermore, once tissue has been excised, it is no longer part of the living organism; oxy and dexoyhemoglobin concentrations begin to diverge from physiologic quantities and if not carefully handled, the hydration of the tissue begins to change.
Photon diffusion theory may be employed to determine optical properties of in vivo samples at source-detector separation longer than five mean-free-paths, where mean-free-path is defined as 1/(μa+μs′). It has been proven to be a not adequate model because boundary conditions and the assumption of multiple scattering in a turbid medium cannot be satisfied. In order to limit interrogation to superficial tissue volumes, such as skin, source-detector separations shorter than five mean-free-paths are more favorable. In-vivo techniques which are capable of measuring optical properties of skin do exist, but have some important limitations. For example, optical properties have been measured of in-vivo skin using visible reflectance spectroscopy with a multi-layer skin model and a genetic optimization algorithm. A multi-layer skin model and a number of fitting parameters, such as layer thickness, chromophores, and scattering properties for each layer, and their corresponding ranges must be chosen carefully in advance to avoid nonuniqueness in the solution space. Some have proposed a model to extract optical properties from diffuse reflectance spectra collected from human skin in-vivo. This technique requires that all of the chromophores contributing to the measured signals are known in advance and the reduced scattering coefficient has a linear relation to the wavelength in order to separate absorption and reduced scattering coefficients from measured reflectance. For the case where all constituent chromophores cannot be determined, the absorption spectra cannot be recovered. In addition, for the case where the reduced scattering coefficient does not have a linear dependence on wavelength, the empirical mathematical model will not recover tissue optical properties properly.
Probes for use in free space or for quantitative measurements of chromophores in tissues that can be reached by an endoscope or similar instrument in which the source and detector are in relatively close proximity with one another has been a significant challenge for quantitative optical methods.
Diffuse optical spectroscopy using frequency modulated light has been employed for years to quantify in-vivo tissue constituents and optical properties. Diffusion approximation to the equation of radiative transport provides a modeling framework for this approach, and gives an accurate description of light propagation in thick tissues as long as detected photons have undergone at least 10 scattering events before they reach the detector. Similarly, this approach is constrained to situations in which the reduced scattering coefficient, μs′ is greater (by an order of magnitude) than the absorption coefficient. In practical terms, this limits the technique to source-detector separations of about 5 mm (depths of about 2.5 mm), wavelengths between 650-1000 nm and modulation frequencies between 50 and 600 MHz.
As source detector separation is reduced to distances smaller than 10 mm, the validity of diffusion approximation is reduced along with ability to accurately recover optical properties and chromophore concentrations. As this distance becomes smaller, the average number of scattering events that photons experience before detection is also reduced. Similarly as one moves to more highly absorbing spectral domains (shorter wavelengths than 650 nm and longer wavelengths than 1000 nm), a reduction in source-detector separation is necessary in order to collect light with reasonable signal to noise ratio. In each of these cases, a simple application of diffusion approximation based modeling will yield inaccurate tissue optical properties and chromophore concentrations.
The problem of quantifying superficial chromophores and optical properties has been solved in the past primarily using multivariate calibration techniques such as the method of Partial Least Squares (PLS). In such an approach, signals are acquired from a set of samples that are representative of the sample of interest. The concentration of the analyte of interest must be known for each sample included in the calibration. By sampling many “reference” samples, an empirical model relating spectral shapes to analyte concentration can be developed. The problem with this approach is that the calibration samples have to be very similar to the target (unknown) sample set of interest. In addition, there has to be a way of recovering the true concentration of the analyte of interest in each of those samples, using a separate method, so that a correlative model can be developed. Note that initial experiments have involved a highly diffusing layer that is, for all practical purposes, infinite in the X-Y dimensions. Clearly this is impractical when thinking in terms of a probe for interrogating, for example, oral tissues.